The goal of the proposed research is to understand biological energy transduction between membranes using the Gram-negative bacterium E. coli as the model system. In E. coli, cytoplasmic membrane energy is coupled to the active transport of important nutrients (iron-bearing sideropheres and vitamin B12) across the outer membrane. The mechanism of energy transduction remains to be elucidated. Several proteins have been shown genetically to participate in the phenomenon, most likely in the form of energy transduction complexes. Emphasizing TonB protein, the protein most directly involved in energy transduction, we will attempt to characterize the structure, function, and regulation of energy transduction complexes. The specific aims of this project are 1.) to determine the composition and stoichiometries of TonB-dependent energy transduction complexes. 2.) to elucidate the role of chemical modifications in TonB-dependent energy transduction. 3.) to delineate the mechanism of coupling between the cytoplasmic membrane proton-motive force and the energy-requiring processes of the outer membrane. 4.) to characterize the mechanisms of transcriptional regulation of TonB. Research in this area will have important ramifications for understanding energy transduction, protein-protein interactions, and membrane phenomena in general.